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However, successful long\u2010read sequencing experiments demand high concentrations of highly purified DNA of high molecular weight (HMW), which limits the utility of established DNA extraction kits designed for short\u2010read sequencing. The challenges associated with input DNA quality intensify further when working with complex environmental samples of low microbial biomass, which requires new protocols that are tailored to study metagenomes with long\u2010read sequencing. Here, we use human tongue scrapings to benchmark six HMW DNA extraction strategies that are based on commercially available kits, phenol\u2013chloroform (PC) extraction and agarose encasement followed by agarase digestion. A typical end goal of HMW DNA extractions is to obtain the longest possible reads during sequencing, which is often achieved by PC extractions, as demonstrated in sequencing of cultured cells. Yet our analyses that consider overall read\u2010size distribution, assembly performance and the number of circularized elements found in sequencing results suggest that column\u2010based kits with enzyme supplementation, rather than PC methods, may be more appropriate for long\u2010read sequencing of metagenomes.<\/jats:p>","DOI":"10.1111\/1755-0998.13588","type":"journal-article","created":{"date-parts":[[2022,1,24]],"date-time":"2022-01-24T00:04:51Z","timestamp":1642982691000},"page":"1786-1802","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":55,"title":["High molecular weight DNA extraction strategies for long\u2010read sequencing of complex metagenomes"],"prefix":"10.1111","volume":"22","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4933-2896","authenticated-orcid":false,"given":"Florian","family":"Trigodet","sequence":"first","affiliation":[{"name":"Department of Medicine The University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1903-756X","authenticated-orcid":false,"given":"Karen","family":"Lolans","sequence":"additional","affiliation":[{"name":"Department of Medicine The University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8957-9922","authenticated-orcid":false,"given":"Emily","family":"Fogarty","sequence":"additional","affiliation":[{"name":"Committee on Microbiology University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4806-9280","authenticated-orcid":false,"given":"Alon","family":"Shaiber","sequence":"additional","affiliation":[{"name":"BioPhysical Sciences Program The University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0281-326X","authenticated-orcid":false,"given":"Hilary G.","family":"Morrison","sequence":"additional","affiliation":[{"name":"Josephine Bay Paul Center for Comparative Molecular Biology and Evolution Marine Biological Laboratory  Woods Hole Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9456-367X","authenticated-orcid":false,"given":"Luis","family":"Barreiro","sequence":"additional","affiliation":[{"name":"Department of Medicine The University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7427-4424","authenticated-orcid":false,"given":"Bana","family":"Jabri","sequence":"additional","affiliation":[{"name":"Department of Medicine The University of Chicago  Chicago Illinois USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9013-4827","authenticated-orcid":false,"given":"A. 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